Increased bronchial irritability (bronchial hyperreactivity) is a characteristic feature of asthma, and understanding its pathogenesis may provide new insights for better treatment. In guinea pigs with acute O3-induced airway injury, we have found cholinergic hyperreactivity which may be related to lipoxygenase product, possibly leukotriene, elaboration. We now aim to (1) document possible regional differences in cholinergic and leukotriene responsiveness of different airway generations in the guinea pig, (2) investigate the cellular mechanisms by which leukotrienes may cause airway responsiveness in this species, and (3) study in vitro whether muscarinic responsiveness and/or cholinergic neurotransmission of certain airway generations is increased in animals with O3-induced bronchial hyperreactivity. To accomplish Aim 1, we will compare effects of the leukotrienes on (a) smooth muscle preparations from different airway generations and on (b) ASM muscarinic responsiveness to endogenously released (via electrical field stimulation (EFS)) versus exogenous ACh. If leukotriene(s) augment(s) EFS responsiveness, possible pre-synaptic effects will be more directly assessed by measuring (3H)ACh release from intramural cholinergic nerve terminals of airways. To further satisfy Aim 2, potential post-synaptic, electromechanical effects will be assessed by the simultaneous measurement of muscle cell membrane potential and force development in different airway generations upon leukotriene stimulation. Possible effects on extracellular Ca++-dependent excitation-contraction coupling will be evaluated in muscle preparations stimulated by tetraethylammonium. To investigate possible pharmacomechanical leukotriene effects on muscarinic contraction that may be extracellular Ca++-independent, muscle preparations will be studied in zero Ca++, La+++ buffer. During the last 2 years of this project, airways from guinea pigs with O3-induced bronchial hyperreactivity will be evaluated employing the above methods to determine whether muscarinic responsiveness and/or cholinergic neurotransmission of certain airway generations is, in fact, increased in this disorder.